Abstract

The experiment was carried out to determine if inclusion
of cocoa pod husks (CPH) in layer diets will affect laying performance and
egg characteristics. Two hundred and sixteen (216) Bovan Brown {BB} layers
(92 weeks old) were randomly assigned to twelve experimental diets for 12
weeks in a completely randomized design. There were three levels of CPH
inclusion; 0%, 10% and 15%. For each level of CPH, diets were further
sub-divided into four and each portion treated with, i) no enzyme, ii)
phytase only, iii) a commercial enzyme cocktail only and iv) a combination
of both phytase and cocktail. The enzyme cocktail was added at a rate of
200g per tonne of complete feed. The phytase was added at the rate of 250g
per ton of complete feed to give a phytase activity of 500 FTU (Phytase
Units)/kg of complete feed.

Overall, adding CPH did not affect average daily
feed intake (ADFI). Hen day egg production for layers on diets with 0%, 10%
and 15% CPH, with a combination of phytase plus an enzyme cocktail (76.19,
73.81 and 66.34 respectively), was better than that of hens on diets without
enzymes. Adding either phytase, a cocktail enzyme, or a combination of the
two improved egg weight. There were no effects of CPH or enzyme addition on
egg quality characteristics. Cocoa pod husk (up to 15%) plus exogenous
enzymes can effectively be used in layer diets without adversely affecting
production performance or egg quality characteristics.

Key words: egg production, egg quality, enzymes

Introduction

Agriculture is the most dominant sector of the economies
of many developing countries. In Ghana for example, it contributes about
33.6% of the Gross Domestic Product (GDP) and about 75% of export earnings (GSS
2008). The livestock and poultry sub-sector is estimated to contribute about
6% of Agricultural GDP. It provides employment and helps to meet the basic
meat and animal product requirements for the growing Ghanaian population (MoFA
1990). Commercial poultry production, in particular, provides easily
accessible and affordable meat and eggs. About 80% of the world’s population
get most of their basic nutrients like proteins, fats and vitamins from meat
and eggs (FAO 2009).

Feeds and feeding constitute a large proportion of the
total expenditure in the poultry industry especially when ingredients like
grains are used (Teguia and Beynen 2004). Cereals such as maize, rice, wheat
and millet constitute the major part of poultry feed and makes up to 60-70%
(Ravindran and Blair 1993). In many developing countries, most of the cereal
that is grown is for human consumption, and hopefully, surpluses are used
for animal feeding. However, the reality is that whatever is available is
competed for by humans and livestock for food and feed, respectively. This
competition raises the cost of poultry production. A high cost of animal
production, translates into a high cost of animal protein. Reducing this
cost component in animal (particularly poultry) production therefore is one
way of reducing the cost of poultry products. It is, therefore, imperative
to find local agricultural residues and by-products to use as alternative
feed ingredients in poultry production. These by-products should be cheap,
readily available and not be detrimental to the birds’ health and
productivity (Teguia and Beynen 2004).

Various by-products from the beer and milling industries,
and from the cocoa processing industry like defatted cocoa cake (a
by-product of the cocoa fermentation process) and cocoa pod husk (CPH), have
been used to varying degrees of success in monogastric feeds (Obikaonu and
Udedibie, 2006; Adama et al 2007; Husaini et al 2010; Nortey et al 2013a;
Manu-Barfo et. al 2013). Cocoa pod husk is a major by-product from
commercial cocoa farming, (Alemawor et al., 2009) and forms over 70% (w/w)
of the whole matured fruit of cocoa. Ghana grows some of the best cocoa in
the world and its economy depends heavily on this crop. However, once the
beans have been obtained from the pod, the latter is of little value. Some
of this by-product is used for products such as some local soaps, but most
of it is not utilized and left to rot on the land. It can however be
potentially incorporated into layer diets to reduce the maize (main energy
source) requirement. It has high levels of protein, energy and other
nutrients. It is however, high in fibre and has high levels of lignin (14%),
non-starch polysaccharides (NSP) like hemicellulose (11%), cellulose (35%)
and pectin (6%) (Alemawor et al 2009). These nutrients are not readily
available to monogastrics (poultry and pigs) because this class of animals
lack fibre-degrading enzymes needed to hydrolyze NSP (Barrera et al 2004).
Undigested NSP can influence intestinal transit time and increase digesta
viscosity. Both result in inefficient nutrient absorption which ultimately
affects growth and egg laying performance of animals and birds. In addition,
phosphorous (P) in plants and plant products such as CPH is available as
phytate-phosphorous (Humer et al 2014) and is not readily available to
monogastrics because they lack the enzyme phytase which is responsible for
phytate hydrolysis. Even if they do, the quantities are insufficient (Golovan
et al 2001). Hence for efficient use of CPH in monogastric diets, it is
important to include exogenous fibre-degrading and phytase enzymes in such
diets. These enzymes are able to hydrolyse fibre and phytate, improve
nutrient utilization and improve performance (Mroz et al 1994; Liao et al
2005; Nortey et al 2013b). Phytase has also been shown to improve amino acid
and energy utilization (Nortey et al 2007).

In Ghana, except for a few of the major feed mills, the
use of phytases and carbohydrases routinely in feed formulation is not
widespread despite the fact that a large proportion of monogastric
feeds are based on plant material. This can result in the under-utilization
of available feed nutrients and large amounts of undigested nutrients being
excreted into the environment. Ultimately there is a drop in production and
an increase in production costs. However, routine use of fibre-degrading
and carbohydrase enzymes will enable feedmills to take advantage of the vast
and diverse feed resources (including non-conventional and sometimes high
fibre feed ingredients) available in the West African sub-region. Thus the
hypothesis of this study was that CPH together with exogenous enzymes can
effectively be used in laying hen diets without adversely affecting
performance. The objectives of this study were therefore to determine the
effect of exogenous enzyme supplementation in diets with added CPH on ADFI,
hen day egg production, feed conversion efficiency (FCE) and egg quality
characteristics.

Materials and Methods

The trial was carried out at the Livestock and Poultry
Research Centre (LIPREC), College of Basic and Applied Sciences (CBAS) of
the University of Ghana (UG).

Processing of cocoa pod husk

Fresh cocoa (Theobromacacao) pods were
harvested from the cocoa plantations of the Cocoa Research Institution of
Ghana, New-Tafo in the Eastern Region of Ghana. They were cracked open to
remove the cocoa beans together with the placenta. The husks were then
chopped into slices (average sized 2cm) at the Product Development Unit of
the Research Institution. They were dried in the sun for about 24 hours to
reduce the moisture content to about 80%. The pre-dried slices were then
passed through a combination mincer and pelleting machine to produce pellets
(about10-12 mm). The pellets were again dried for about 48-72 hours to
further reduce moisture content to about 10% and stored until use.

Experimental diets

Twelve (12) experimental diets (T1 to T12) were used in
the trial. Three main diets were initially formulated to contain 0%, 10% and
15% CPH respectively. Each main diet was further sub-divided into four
parts. Part one was not treated any further. Parts two, three and four were
treated with phytase alone (300g/tonne of complete feed), a commercial
enzyme cocktail alone (250g/tonne of complete feed), and a combination of
both phytase and a cocktail enzyme at the stated inclusion levels. Thus T1 (CHO),
T2 (CHO-PHY), T3(CHO-EC) and T4 (CHO-PHY-EC) represented diets with a) 0%
CPH with no exogenous enzyme, b) 0% CPH plus phytase alone, c) 0% CPH plus
enzyme cocktail alone, and d) 0% CPH plus a combination of phytase and an
enzyme cocktail. Treatments five to eight (CH10, CH10-PHY, CH10-EC and
CH10-PHY-EC), and nine to twelve (CH15, CH15-PHY, CH15-EC and CH15-PHY-EC)
represented diets with 10% and 15% CPH respectively, and with the same
combination of enzymes as was in T1 to T4. The enzyme cocktail contained
phytase, amylase, protease, cellulase, xylanase, β-glucanase and pectinase
and was supplied by Zoetis under the brand name Enziver.

Experimental birds and design

Two hundred and sixteen (216) Bovan Brown {BB} layers at
92 weeks old were randomly assigned in battery cages to twelve experimental
diets in a Completely Randomized Design for 12 weeks. The experiment was set
up in a 3 x 4 factorial arrangement of treatments (3 levels of CPH x 4
levels of enzyme treatment). There were 18 birds in each treatment and 6
birds per replicate. The laying hens were allowed seven (7) days on the
experimental diets for conditioning. The experiment consisted of three
periods of 28 days each. Birds were fed the same treatment diet during the
experimental period. Water was provided ad libitum. A known
amount of feed was provided every morning, and feed left over after 24 hours
was collected and weighed to determine feed disappearance. This amount
represented the average daily feed intake (ADFI). Feed conversion efficiency
(FCE) was calculated as the ratio of weight of eggs to feed intake. Eggs
were collected twice a day at 08:00 and 16:00 and grouped according to
treatment and replicate. Records of egg weight, hen-day and hen-housed
egg production performance, were kept daily and summarized on a weekly
basis.

Chemical Analysis

The proximate chemical composition of all the major
ingredients used was analyzed using methods outlined in AOAC (1995). Calcium
and phosphorus were determined according to the methods outlined by James
(1996) and AOAC (1995).

Egg quality analysis

External and internal egg quality measurements were
determined on days 28, 56 and 84. Two eggs laid within a two hour period
were selected from each replicate group. Parameters measured included shell
thickness, egg length and width and albumen height. These measurements were
taken within 24 hours after collection and at room temperature. A digital
caliper was used to measure egg width (at the broadest end) and length. Egg
weight was measured with an electronic balance (OHAUS-Pioneer ™, Ohaus
Corp., USA) with a sensitivity of 0.01g. The egg was then cracked and shell
thickness and albumin height determined using a digital caliper (A & D
Company Ltd). The height of the thick albumen was measured between the yolk
and the edge of thick albumen. Three points were measured and the average
taken. The Haugh Unit was calculated as has been described by Haugh (1937):

Where:

HU = Haugh unit

H= observed height of the albumen in millimeters

W = weight of egg in grams

G = the gravitational constant, 32.2

Statistical Analysis

All data gathered were subjected to statistical analysis
using the Generalised Linear Model procedure of the Statistical Analysis
Systems Institute (SAS 1999). Significant differences among means were
separated using the Student Newman-Kuels (SNK) Test. The results from
the different breeds were handled separately.

A priori, it was decided
to compare the following treatments which were of particular interest, using
contrasts:

All diets without enzymes versus all diets with both
phytase plus cocktail

All diets without enzymes versus all diets with only
a cocktail

All diets without enzymes versus all diets with only
phytase

All diets with only phytase versus all diets with
only a cocktail

All diets with phytase alone versus all diets with
both phytase plus a cocktail

All diets with only a cocktail versus all diets with
both phytase and a cocktail

Results

Table 1a shows the chemical composition of CPH. The diets
used in the experiment, showing CPH levels and combination of enzymes (T1 –
T12) are shown schematically in Table 1b. The composition and calculated
values of the layer diets are shown in Table 2. Protein levels dropped from
16.24 to 15.34 as the level of wheat bran reduced and CPH increased. This
was due to the relatively lower protein content of CPH compared to wheat
bran. A higher fibre content of CPH also meant that the levels of total
dietary fibre increased as the level of CPH also increased.

Table 1a:
Chemical composition of cocoa pod husk

Parameter

Concentration (%)

Dry matter

85.7

Crude protein

7.04

Crude fibre

31.1

Total ash

9.6

Ether extract

5.93

Calcium

0.81

Phosphorous

0.44

Production parameters

Average
Daily Feed Intake

For birds on 0% CPH, those on CH0 and CH0-PHY recorded
the minimum and maximum ADFI respectively. For diets with a cocktail enzyme
alone (CH0-Phy), ADFI was not different from what was obtained for birds fed
a diet with only phytase. At the 10% CPH inclusion, a similar trend was
observed, where birds on diets with no enzyme ate less than those on diets
with enzymes. However for birds on CH10-PHY, ADFI was lower than for birds
on CH10-PHY-EC. Similarly, at the 15% of level of CPH inclusion, the trend
in ADFI as was observed at lower inclusion levels, was observed here as
well. Birds on CH15 had an ADFI which was lower than the ADFI of birds on
CH15-PHY-EC but similar to that of birds on the remaining two diets.

Hen day egg production

Birds on CH0-PHY-EC and CH10-PHY-EC had hen day egg
production values which were similar, but higher than egg production values
of hens on the remaining treatments. For the rest of the treatments with 0%
CPH, birds on the first three treatments had similar egg production values.
On the average birds on diets with 15% CPH recorded lower egg production
values compared to those on either 0 or 10% CPH.

Egg weight

For each level of CPH inclusion (0, 10 and 15%) egg
weights seemed to increase with enzyme supplementation. At the 0% CPH
inclusion, egg weights for birds on CH0-PHY-EC was similar to those on
CH0-PHY, but heavier than those on the remaining two. At 10% CPH inclusion,
birds on CH10-PHY-EC had the heaviest eggs which was heavier than that for
birds on 10CP-Cont and similar to the remaining two.

Average daily feed intake of, birds on treatments without
any added enzyme was lower than that of birds with a combination of both
enzymes. Birds fed on diets with either phytase, or a cocktail alone
irrespective of CPH inclusion level, ate less than birds that were fed diets
with a combination of phytase and a cocktail.

Enzyme addition

Average hen day egg production of birds on diets without
any enzyme irrespective of CPH inclusion was 60.6% and was lower than egg
production of birds on both phytase plus an enzyme cocktail (72.11%). Adding
a cocktail enzyme resulted in an egg production of 65.1%, which was better
than adding only phytase to the diet (64.32%). The trends noted for hen day
production were observed in egg weights as well. Egg weight of birds on
diets without added enzyme irrespective of CPH inclusion (61.15g) was lower
than egg weight of birds that had been fed diets containing both phytase
plus an enzyme cocktail (63.72g). Compared to eggs from diets that had
phytase alone (63.05g), birds fed diets with a cocktail enzyme laid eggs
that averaged 63.24g in weight, and these were different from one another.

A. priori contrasts were performed to determine which of
the enzymes and/or combination of enzymes resulted in improved production
characteristics irrespective of CPH inclusion. A comparison between
diets with added cocktail enzymes or those with only phytase, versus diets
with no enzymes at all indicated that the birds did not benefit
significantly (Table 5). However, comparing the efficacy of the two classes of enzymes
showed that improvements occurred only when both enzymes were added to the
diet and the results compared to diets without any added enzyme.A similar
trend was observed in hen day egg production and egg weight, where the birds
reacted favourably only when a combination of the two enzymes was added to
the diet. Comparing the two enzymes separately however showed that the
cocktail enzyme performed better than phytase.

Effect of CPH

There was similarity in ADFI in spite of increasing fibre
levels and slightly reducing energy densities as the level of CPH increased
in the diet. Generally where gut fill is not a factor, monogastrics like
birds will eat more of a diet that is low in nutrient density, in an attempt
to meet their daily nutrient requirements, particularly energy (Leeson and
Summers 1997). This phenomenon has been observed in broilers by Alemawor et
al (2010) and in layers by Umar Faruk et al (2010). However since it
was not observed in the current study, it may be concluded that dietary
energy dilution was not enough to elicit an increased feed intake. The
layers used in this trial were 92 weeks old and thus probably had well
developed caeca with a great number and diversity of microbes which were
capable of breaking down fibre and releasing nutrients in the form of
volatile fatty acids (VFA). This will contribute to the maintenance energy
requirements of the birds and prevent any expected increase in ADFI.

Hen housed egg production generally decreased with
increasing levels of CPH. There was a 2 percentage point drop in egg
production when the level of CPH in the diet went from zero to 10%, whilst a
10% dip in production occurred when the CPH level was further increased to
15%. It can be assumed from this observation that any ANF present in CPH
exerted a negative effect on egg production at levels beyond 10%.

There were similar egg weights for diets with 0, 10, and
15% CPH respectively. Among the factors affecting the weight of an egg ADFI
plays a key role. Thus similarities in ADFI over the three levels of CPH
inclusion, could explain the lack of differences in egg weight that were
observed. Feed conversion efficiency in egg production is the efficiency of
converting feed into eggs. With the observed trend in ADFI and egg weights
the lack of effect of CPH level in the diet on FCE was expected. The lack of
an effect of CPH on shell thickness may be due to the fact that the levels
of both calcium and P in the diet, which are the main minerals needed for
egg shell formation were adequate and not out of balance. Internal egg
parameters were not affected by the level of CPH in the diet.

Effect of enzyme addition

Enzymes are biological catalysts that speed up chemical
reactions in biological systems. Although addition of phytase or a cocktail
enzyme by themselves tended to increase ADFI in this trial, a combination of
both phytase and a cocktail, caused a more drastic effect. This trend was
the reverse of what was observed with pigs (Nortey et al 2013b; Kies et al
2005; Madrid et al 2013) and in poultry (Osei and Oduro 2000) where enzyme
addition tended to reduce ADFI.

Table
5: p-values of A priori treatment comparisons of interest:
production parameters

The theory behind a drop in ADFI upon enzyme supplementation
is the phenomena of nutrient uplift. It is generally assumed that exogenous
enzyme addition causes a release of nutrients. This will cause the daily
nutrient requirement of animals to be met faster. In a situation like this, some
mechanisms that control feed intake including the glucostatic, thermostatic,
lipostatic, aminostatic and ionostatic theories will come into play to stop
further feed ingestion. However, the glucostatic theory of feed intake
regulation as observed in some mammals is said to be either not present in
poultry or is not well understood (Richardson 1970; Smith and Bright-Taylor
1974). In the absence of such a theory occurring in this trial, one can
speculate that inclusion of enzymes, in addition to releasing nutrients, may
have reduced (or prevented) any possible fibre-inducing intestinal viscosity.
This will result in slightly faster intestinal transit times leading to a
feeling of emptiness and hence the need to increase ADFI. The fact that a
combination of both enzymes in the diet resulted in greater ADFI compared to
either enzyme alone indicates that there is synergy between both enzymes. Hen
day average egg production and egg weights also increased with added enzymes at
each level of CPH inclusion. This could be a direct result of the increased ADFI.
Some factors that influence egg production and weight are level of protein and
balance of amino acids, fat, and water intake. For diets that contained CPH,
levels of these nutrients tended to reduce slightly as a result of the dilution
effect of increased fibre from the CPH. However since hen day production and egg
weights tended to improve, it can be speculated that enzyme addition improved
nutrient availability.

The observed results in the a priori comparisons were
expected since in addition to phytase, the cocktail enzymes contained other
enzymes which together would act synergistically to improve nutrient
digestibility (Nortey et al 2008).

Conclusions

Cocoa pod husk was included in diets for layers up to a
level of 15% and in the presence of enzymes without affecting production.

There was a synergistic effect between phytase and a
cocktail enzyme in improving egg laying and production performance.